Cause

Measuring only about 20 nm in diameter, circoviruses are comparatively small, nonenveloped, icosahedral viruses that contain relatively simple, circular, single-stranded DNA genomes, making them among the smallest and simplest pathogens so far known. Their small genomes of approximately 2 kb encode for only two major proteins, the structural capsid (Cap) protein and a replicase (Rep) protein. Compared with budgerigar fledgling disease polyomavirus and other DNA viruses, they are genetically diverse, even within species. Despite this, little association has been demonstrated between genotypes and pathotypes of any of the members of the Circoviridae.

Clinical Signs

PBFD generally affects juvenile or young adult psittacine birds but all ages may succumb to the disease. Two syndromes are recognized, an acute form, which occurs in nestlings and African grey parrots (Psittacus erithacus), and a chronic form, which occurs in many species of psittacine birds. In acute disease, there is rapid development of depression associated with leucopenia, green diarrhea, and death caused by hepatic necrosis. High titers of BFDV may be detected in the liver and bile of affected birds and some may die of liver failure without obvious feather lesions. African grey parrots often die within 1 week of the development of clinical signs and, depending on the age of the nestling, many diseased contour feathers may be shed all at once or only the primary flight feathers may be affected. Feather necrosis causing fractures of the developing calamus and accompanying intrapulp hemorrhage are the predominant clinical findings. Affected feathers fracture from the point of necrosis, usually before the feather has unsheathed. Feather tracts may become acutely swollen and sensitive because of inflammation and edema, and the birds often become systemically ill, anorexic, and/or regurgitate food. Death may occur suddenly within 1 to 2 weeks of developing clinical signs.

More commonly, PBFD develops as a chronic disease that is insidious in its development and progression; dystrophic feathers replace normal ones as they are molted. In this manner, a PBFD-affected bird may gradually lose its plumage without other signs of illness (Fig. 39-1). The pattern of feather dystrophy is related to the stage of molt that the bird is in when the disease first begins; it is usually bilaterally symmetrical and slowly progressive. Dystrophic feathers are usually short and have one or more of the following characteristics: fault lines across the vanes; a thickened or retained feather sheath; blood within the calamus; an annular constriction of the calamus; and curling (Fig. 39-2).

Figure 39-1 Chronic PBFD in a yellow crowned kakariki (Cyanoramphus auriceps) demonstrating generalized plumage deficits caused by bilaterally symmetrical feather dystrophy.

Figure 39-2 PBFD-affected feather demonstrating annular constriction of the developing calamus (arrow) and thickening of the feather sheath (star).

In cockatoos, the powder-down feathers, or pulviplumes, are often the first feathers affected. PBFD-affected pulviplumes are fragile or develop an abnormally thickened outer sheath that fails to disintegrate. Pulviplume follicles may atrophy and thus create bare powder-down patches. Arrested production of powder down causes the plumage to become dull and the beak to become glossy. Claw abnormalities occur occasionally, and generally develop well after feather and beak lesions become apparent. Feather loss is symmetrical, usually affecting the powder-down feathers first in cockatoos, and then flight and tail feathers, before progressively involving the rest of the body until birds appear to be bald. The pattern of feather loss is dependent on the stage of molt that the bird is in when clinical signs commence. Affected developing feathers fracture at an annular constriction, typically near the calamus. The beak becomes progressively elongated, develops fracture lines, and may eventually fracture or slough off. Chronically affected birds are predisposed to secondary infections, presumably because of immunosuppression.

On the extremities, PBFD-induced hyperkeratosis may cause the skin to appear excessively scaly or it may be thickened and moist. Sunlight-exposed skin may become darkly pigmented. Chronic skin ulcers may occur at the elbows and wing tips. Beak, and less commonly, claw deformities occur in some PBFD-affected birds, particularly in cockatoos. The beak may become abnormally soft and brittle and the upper and lower tips elongated. Transverse or longitudinal fractures or delaminations often occur. In severe cases, necrosis of the oral epithelium and osteomyelitis may cause the beak to slough. Secondary disease problems commonly exist. These include cryptosporidiosis and bacterial, mycotic, and other viral infections. Most birds with chronic disease eventually have difficulty eating, lose weight, and die.

In smaller grass parrots such as Psephotus and Neophema spp., apparently normal feathers which fall out or are effortlessly plucked, may be the only clinical sign. The first clinical sign in birds with green plumage may be the development of yellow feathers, which may appear normal in other respects.

Diagnosis

PBFD has distinct pathologic features and, although in most circumstances a presumptive diagnosis may be achieved by clinical examination alone, further tests must be done to differentiate PBFD from similar diseases of the integument (e.g., acute avian polyomavirus infection, chronic hypothyroidism). Clinicopathologic derangements such as anemia and hypoproteinemia are nonspecific. Histologic examination of feather follicle or feather biopsies may confirm clinical disease but is not suitable for diagnosing incubating BFDV infection or detecting BFDV carriers. Necrosis of feather epidermal cells and the presence of relatively large intracytoplasmic inclusions in macrophages and keratinocytes is characteristic of the disease. Similar viral inclusions occur in the bursa of Fabricius. Lesions are usually absent in the apterial and interfollicular skin, although BFDV antigen may be found here by immunohistochemistry. This may also be used to detect antigen in the bursa of Fabricius, spleen, pharynx, crop, esophageal epithelium, and other tissues.

Unlike other members of the Circovirus genus, BFDV is a hemagglutinating virus and has been shown to agglutinate erythrocytes from guinea pigs, geese, and many species of psittacine birds. To date, no method has been described to cultivate BFDV in vitro successfully. Thus, research into the biologic characteristics, pathophysiology, and mode of replication has required the use of virus purified from the tissues of infected birds or recombinant proteins.

Virus detection by hemagglutination (HA) assay is a sensitive diagnostic test for detecting BFDV antigen emission in feather material. Chronically affected birds typically excrete very high titers (>1 : 40,960), often in the absence of detectable antibodies measurable by hemagglutination inhibition (HI) assay.¹⁰ HA may be used to detect virus excretion in feathers, liver, bile, and feces and may be performed on actively growing feather pulp or dry keratinized feathers.

Virus DNA detection by polymerase chain reaction (PCR) assay is also available as a diagnostic test and may detect BFDV infection in circulating leucocytes or feather material. The results must be interpreted in relation to the clinical signs and results of other tests, because cross-contamination caused by unclean collection and laboratory technique is possible.^4,10,12

Serology may be done by HI, enzyme-linked immunosorbent assay (ELISA), or blocking ELISA (bELISA) and is useful for detecting BFDV-infected flocks or seroconversion in individual birds.^8,10,24 A high plasma HI antibody titer (>1:320) in an adult bird is a good indicator that it does not have chronic PBFD. Nestlings with incubating infection or acute disease may have low and declining antibody titers. The HI test may be done on serum, plasma, or whole blood collected on filter paper.

Pathogenesis and Epidemiology

The incubation period of PBFD may be as short as 21 days but it is probably dependent on the dose of virus, age of the bird, stage of feather development, and absence of immunity. Primary virus replication probably occurs in the bursa of Fabricius and/or gastrointestinal tract lymphoid tissue. Secondary virus replication occurs in the liver and thymus, and probably in other tissues. The target organ is the epidermis and the manifestation of skin disease requires a molt. Consequently, birds that become infected after feather development has completed may not develop clinical signs until their next molt. This could take 6 months or longer. Most birds that succumb to PBFD are younger than 2 years. However, all age groups should be considered susceptible to circovirus infection. Long-term exposure and/or stress are probably required for infection and seroconversion in adult birds.

Very little is known about the steps involved in the replication of BFDV but it is possible that it uses a similar mechanism for cellular entry into macrophages as that discovered for porcine circovirus The capsid protein contains a nuclear localization signal that assists viral entry into the nucleus,⁷ but the genome may also be able to enter the nucleus during mitosis by associating itself with cellular chromatin. Circoviruses are highly dependent on cellular enzymes for their replication via the synthesis of a complementary strand to form a double-stranded DNA (dsDNA) replicative form and then by rolling circle replication. How assembled virions are transported into the nucleus to form the characteristic paracrystalline arrays, which are readily seen histologically, is not precisely known, but BFDV Cap and Rep are known to interact,⁷ which may facilitate transport of encapsidated virions into the cytoplasm.

BFDV is epitheliotrophic and primary sites of replication include the skin (including the epithelium of the beak and claws), liver, gastrointestinal tract, and bursa of Fabricius. The virus replicates to high titers in these tissues and also spreads to the spleen, thyroid, parathyroid, and bone marrow. Skin lesions are associated with apoptosis of keratinocytes.

The maximum incubation period of PBFD is unknown, because the appearance of clinical signs depends on the stage of molt of the bird; however, experimentally infected nestlings may show signs of acute disease from 21 to 28 days postinfection. BFDV is excreted via the feathers and the gastrointestinal tract. Consequently, high concentrations of BFDV antigen may be detected in liver tissue, bile, crop secretions, feces, and feather dander. Horizontal transmission by oral and/or intracloacal ingestion of virus excreted via feces, feather dander, and crop secretions is likely the primary mechanism of viral spread. Experimental infection may also be accomplished by intramuscular administration of the virus. Vertical transmission is suspected, because BFDV DNA has been amplified from embryonated eggs by PCR assay.¹⁸

All species of psittacine birds should be considered to be susceptible to BFDV. Depending on the species and age, not all birds infected with BFDV progress to develop clinical signs of disease. Spontaneous clinical recovery from acute PBFD may occur rarely in many species, including budgerigars (Melopsittacus undulatus), lorikeets, and Eclectus and Agapornis spp. but these birds typically become carriers. However, most chronically affected birds do not recover from the disease. Individual lorikeets and eclectus parrots may develop protective HA antibody with intermittent cessation of virus excretion. In contrast, others, such as African grey parrots and black cockatoos (Calyptorhynchus and Callocephalon spp.), appear highly susceptible and may die before clinical signs of feather loss develop. In contrast, cockatiels seem to be resistant to BFDV infection and subsequent PBFD.²³ White cockatoos (Cacatua spp.) typically develop the chronic form of the disease, with high HA titers detectable in feathers and feces and no detectable HI titer. The reasons for the variations in clinical disease remain to be explained.

Compared with APV, BFDV is a relatively genetically diverse species, with BFDV nucleotide sequence similarities reported to range from 84% to 99%. Based on phylogenetic analyses and differences in pathogenicity, the existence of strains of BFDV has been proposed but this has yet to be confirmed experimentally. Individual serotypes were not found in the one cross-reactivity study¹⁰ undertaken to date. However, studies on BFDV isolates from PBFD-affected cockatiels (Nymphicus hollandicus) have shown some degree of serologic and phylogenetic divergence, but this is insufficient to justify classifying these as distinct strains or species.²³

In infected flocks viral DNA may be detected in up to 28.0% of birds⁶ in wild flocks and up to 83% to 90% of birds in captive flocks.¹⁰ Seroprevalences up to 62% in captive flocks and up to 94% in wild flocks have been reported. In wild birds, PBFD has been confirmed in most Australian psittacine bird species, as well as in parrots and cockatoos throughout Indonesia, Papua New Guinea, and New Zealand. In Australia, flocks of wild cockatoos may have a disease prevalence of 20% and a seroprevalence of 60% to 80% and infection is probably maintained in a population by diseased birds and contaminated nest hollows. Epidemics may occur in susceptible wild or aviary flocks. Virus transmission is probably predominantly by horizontal spread, but carrier birds may contribute by vertical transmission. Virus infectivity probably persists in contaminated nests for many months or years. Aviary flocks with a history of PBFD usually have a high seroprevalence. In these situations, PBFD-affected birds are often the progeny of hens with low or nondetectable serum antibody levels.